Page 119 - Multidimensional Chromatography

Coupled-Column Liquid Chromatography 111

Table 5.1 Comparison of off- and on-line multidimensional LC techniques
Off-line multidimentional chromatography
Advantages Disadvantages
• Easy to carry out by collection of • Labour intensive
column effluent
• Can concentrate trace solutes from • More time consuming
large volumes
• Can work with two LC modes that • Sample loss or contamination
use incompatible solvents during handling
On-line multidimentional chromatography
Advantages Disadvantages
• Easy to automate • Incompatibility of different mobile
phase system
• No loss or contamination • Separation obtained on the first
column can, at least partly, be
reduced on the second column
• Decreased total analysis time • Requires automated or semi-automated
instrumentation
• More reproducible

columns, mobile phase composition, the nature and number of analytes, and the type
of matrix and its related interferences. In order to achieve this, simple valving cir-
cuits can be used with conventional LC apparatus.

5.2 THEORETICAL ASPECTS

In order to optimize separations on coupled-column liquid chromatographic systems
under the conditions of solvent modulation, we need to consider the parameters
which affect the resolution, as follows.
The basic measure of the efficacy of a single-column chromatographic system in
separating two neighbouring peaks can be effected by the resolution (R s ), which is
equal to the ratio between the two peak maxima, t r (distance between the peak cen-
tres) and the average base width of the two peaks, as follows (3):

2(t r,2 t r,1 ) t r
t r
R S (5.1)
w (w 1 w 2 ) 4 t
where t r,1 and t r,2 are the retention times, w 1 and w 2 are the peak widths measured by
the baseline intercept and t is the mean standard deviation of a Gaussian peak.
For closely spaced peaks, the resolution may be expressed as the product of three
factors (a,b,c), which are related to the adjustable variables of a chromatographic
system as follows:
1 k 2 1/2
N
R S 1 k 2 16 (5.2)
(a) (b) (c)